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Biomedical subjects

M R Young

Publications and source records attributed to M R Young.

At least 109 records · Page 6Linked to original sources
PubMed1993

Stimulation of splenic T-lymphocyte function by endogenous serotonin and by low-dose exogenous serotonin.

The modulatory effects of endogenous serotonin on splenic T-cell activity were investigated using two distinct approaches. The first approach showed that pretreatment of mice with p-cholorphenylalanine (PCPA) to deplete intracellular stores of serotonin reduced the capacity of their splenic T cells to proliferate and to express interleukin-2 receptor (IL-2R) in response to concanavalin A (Con A). These responses could be restored by the addition of serotonin to the spleen cell cultures. In contrast, PCPA treatment did not effect stimulation of spleen cells to produce IL-2. The second approach showed that T-cell proliferation to Con A as well as to IL-2 was diminished by the presence of antagonists to the serotonin-2 receptor (5-HT2R). The effects of low doses (100 ng/ml) of exogenously added serotonin on functions of normal spleen cells were also examined. At this low dose, serotonin stimulated splenic T-cell proliferation in response to IL-2, and enhanced both proliferation and IL-2 production in response to a suboptimal concentration of Con A. These results show autologous serotonin to be required for T-cell activation and that the activation of suboptimally stimulated T cells can be augmented with low doses of exogenously added serotonin. These data also suggest that the positive regulation of T-cell function by serotonin is mediated through 5-HT2R.

Animals↗
PubMed1993

J5 prevents endotoxin shock in suckling rats.

A sublethal dose of endotoxin (lipopolysaccharide: LPS) can induce endotoxin tolerance. It is not well known whether LPS feeding causes endotoxemia and endotoxin tolerance in the newborn. Since Rc mutant Escherichia coli LPS (J5) does not have toxic or lethal effects, J5 is used to induce endotoxin tolerance in this study. This study showed that both an intraperitoneal (ip) injection and feeding of Rc mutant Escherichia coli LPS (J5) induced endotoxin tolerance in suckling rats.

Administration, Oral↗
PubMed1992

Myelopoiesis-associated immune suppressor cells in mice bearing metastatic Lewis lung carcinoma tumors: gamma interferon plus tumor necrosis factor alpha synergistically reduces immune suppressor and tumor growth-promoting activities of bone marrow cells and diminishes tumor recurrence and metastasis.

Metastatic Lewis lung carcinoma (LLC) tumors stimulate myelopoiesis and, consequently, induce bone marrow cells to become immune suppressive to T cell blastogenesis and macrophage activation for tumor necrosis factor alpha (TNF-alpha) secretion. The suppressor cells phenotypically resembled granulocytic-monocytic progenitor cells. In order to diminish the presence of these immune suppressor cells, LLC-bearing mice were treated with low doses of gamma interferon (IFN-gamma) (100 units/mouse) plus TNF-alpha (10 units/mouse). Treatment of LLC-bearing mice with these low doses of IFN-gamma plus TNF-alpha diminished the suppressive activity of their bone marrow cells, as measured by the effect on normal macrophage activation to secrete TNF-alpha. In in vivo adoptive transfer studies, bone marrow from placebo-treated LLC-bearers stimulated tumor establishment and metastasis, while the bone marrow of IFN-gamma-plus TNF-alpha-treated tumor-bearers diminished LLC establishment and metastasis. The effect of the low dose treatments with IFN-gamma and/or TNF-alpha on the recurrence of excised s.c. tumors was also assessed. Treatment of mice following tumor excision with either IFN-gamma, TNF-alpha, or the combination of IFN-gamma plus TNF-alpha reduced recurrence. However, in the animals with recurring tumors only the combined IFN-gamma plus TNF-alpha treatment effectively diminished the development of lung metastases. These results demonstrate that low dose IFN-gamma plus TNF-alpha treatment diminishes the presence of suppressor and tumor growth-promoting activities of bone marrow and reduces tumor recurrence and metastasis.

Animals↗
PubMed1992

Stimulation of the metastatic properties of Lewis-lung-carcinoma cells by autologous granulocyte-macrophage colony-stimulating factor.

Using both polymerase-chain-reaction analysis and the soft-agar colony-forming unit assay, granulocyte-macrophage colony-stimulating factor (GM-CSF) was shown to be expressed by cloned metastatic Lewis-lung-carcinoma (LLC-LN7) cells but not by non-metastatic LLC-C8 cells. Furthermore, the metastatic LLC-LN7 cells were shown to respond both to autologous GM-CSF and to exogenous recombinant GM-CSF (rGM-CSF). In the presence of neutralizing anti-GM-CSF antibodies, the metastatic LLC cells became less able to migrate or to adhere and invade through a reconstituted basement membrane. Moreover, the addition of rGM-CSF further enhanced the capacity of the metastatic LLC cells to adhere to the reconstituted basement membrane. This stimulation of metastatic properties of the LLC cells by either autologous or exogenous GM-CSF was associated with enhanced endogenous protein phosphorylation. Two proteins of approximately Mr 45,000 and Mr 64,000 were the dominant target proteins to be phosphorylated by the presence of GM-CSF. These results suggest that autologous GM-CSF may function as an autocrine stimulator of the metastatic properties of metastatic LLC cells.

Cell Movement↗
PubMed1992

Amalgamation of two endocytic probe techniques: fluoresceinated cationized ferritin can show up, sequentially, selected structures, first in living cells and then by electron microscopy.

Fluorescein isothiocyanate-cationized ferritin (FITC-CF) has hitherto been used mainly to identify structures in living cells by light microscopy, by virtue of its fluorescent properties. We show here that this conjugate can be used, after immediate fixation of the same cell sample and preparation of thin sections, to recognise the same structures, by virtue of the ferritin's electron opacity. The conjugate should thus have a new use as a single-application, dual-purpose probe, e.g. in endocytic studies. The procedure may have advantages over similar dual-purpose probes in not requiring staining or special treatment.

Animals↗
PubMed1992

Tumor-derived cytokines induce bone marrow suppressor cells that mediate immunosuppression through transforming growth factor beta.

Normal bone marrow cells become immunosuppressive when cultured with supernatants of metastatic Lewis lung carcinoma (LLC-LN7) cells. The suppressor-inducing activities in the LLC-LN7 supernatants are interleukin-3 and granulocyte/macrophage-colony-stimulating factor. In the present study, the mechanisms by which these induced suppressor cells (LLCsup-BM) mediate their immunosuppression were investigated. The suppression by LLCsup-BM of splenic concanavalin CA blastogenesis was not dependent on cell contact since immunosuppression occurred regardless of whether the LLCsup-BM were separated from the responder spleen cells by a permeable membrane or if the LLCsup-BM were cocultured with the spleen cells. Culture supernatants of LLCsup-BM also inhibited T cell blastogenesis, being more suppressive than were supernatants of control bone marrow cells, which had been precultured with medium. The suppression by the soluble inhibitors elaborated from the LLCsup-BM was not restricted to the inhibition of T cell function as the supernatants also inhibited the natural killer activity of normal spleen cells. Studies to determine the identity of the suppressive activity produced by the LLCsup-BM showed increased levels of transforming growth factor beta (TGF beta) in their supernatants. Immunosuppressive bone marrow and spleen cells obtained from mice bearing metastatic LLC-LN7 tumors also secreted more TGF beta than did the cells obtained from normal mice. When anti-TGF beta antibodies were added to the LLCsup-BM supernatants, the suppressive activity was diminished. These results suggest that the LLCsup-BM mediate at least part of their immunosuppression through production of TGF beta.

Animals↗
PubMed1992

Regulation of murine T-lymphocyte function by spleen cell-derived and exogenous serotonin.

The modulatory effects of serotonin on T-cell activity were investigated. T-cell blastogenesis of normal spleen cells was slightly stimulated by the addition of low doses (1 and 10 ng/ml) of the inducer of serotonin release, fenfluramine. In contrast to the stimulatory effects of low doses of fenfluramine, high doses of fenfluramine (1 and 10 ug/ml) or of exogenously added serotonin (> or = 0.1 ug/ml) inhibited T-cell activation. Both the stimulation by low dose fenfluramine and the inhibition by high dose fenfluramine were accentuated by pretreating mice with tryptophan to heighten intracellular stores of serotonin and then inducing serotonin release. Pretreatment of mice with the serotonin inhibitor p-chlorophenylalanine (PCPA) abolished the fenfluramine inhibition of T-cell activation indicating that the fenfluramine inhibitory effect was mediated via endogenous spleen cell-derived serotonin. However, the PCPA treatment diminished T-cell activation. These results suggest that endogenous serotonin causes a biphasic dose-response effect on T-cell activity with serotonin being required for optimal T-cell function, low doses being immune stimulatory and higher doses being suppressive.

Adjuvants, Immunologic↗
PubMed1992

Activation of protein kinase A increases the in vitro invasion, migration, and adherence to reconstituted basement membrane by Lewis lung carcinoma tumor cells.

Prostaglandin E2 (PGE2) stimulated in vitro invasion, migration, and adherence to reconstituted basement membrane by metastatic Lewis lung carcinoma (LLC) clones, but this stimulation was blocked by protein kinase A (PKA) inhibitors and by expression of a transfected mutant RI alpha which blocks PKA activation. In vitro migration and adherence of metastatic LLC transfectants was heightened by expression of a transfected C alpha gene and further heightened by Zn2+ induction of the expression vector. Nonmetastatic LLC were not migratory nor invasive. However, nonmetastatic LLC that were stably transfected with a C alpha gene were both migratory and invasive, particularly when C alpha expression was further induced with Zn2+. The results of these in vitro studies show that PKA can enhance the metastatic characteristics of LLC.

Animals↗
PubMed1991

Ammonium chloride, an inhibitor of phagosome-lysosome fusion in macrophages, concurrently induces phagosome-endosome fusion, and opens a novel pathway: studies of a pathogenic mycobacterium and a nonpathogenic yeast.

The weak base ammonium chloride has been previously reported to inhibit lysosomal movements and phagosome-lysosome (Ph-L) fusion in cultured mouse macrophages (M phi), thus reducing delivery, to an intraphagosomal infection, of endocytosed solutes that have concentrated in secondary lysosomes. We have now addressed the question, whether NH4Cl might affect any direct interaction (if it exists) between such infection phagosomes and earlier, nonlysosomal compartments of the endocytic pathway, i.e., solute-containing endosomes. The phagosomes studied were formed after ingestion of the mouse pathogen Mycobacterium microti and the nonpathogenic yeast Saccharomyces cerevisiae; and the endosomes were formed after nonreceptor-mediated endocytosis of electronopaque and fluorescent soluble markers. By electron microscopy, survey of the cell profiles of M phi that had been treated with 10 mM NH4Cl so that Ph-L fusion was prevented, and that displayed many ferritin-labeled endosomes, revealed numerous examples of the fusion of electronlucent endosomes, revealed numerous examples of the fusion of electronlucent vesicles with phagosomes, whether containing M. microti bacilli or S. cerevisiae yeasts. Fusion was recognized by transfer of label and by morphological evidence of fusion in progress. The fusing vesicles were classed as endosomes, not NH4Cl-lysosomes, by their appearance and provenance, and because lysosome participation was excluded by the concurrent, NH4Cl-caused block of Ph-L fusion and associated lysosomal stasis. No evidence of such phagosome-endosome (Ph-E) fusion was observed in profiles from M phi treated with chloroquine, nor in those from normal, untreated M phi. NH4Cl-treated living M phi that had ingested yeasts at 37 degrees C, followed by endocytosis of lucifer yellow at 17 degrees C (to accumulate labeled endosomes and postpone label passing to lysosomes), were then restored to 37 degrees C. Fluorescence microscopy showed that as many as half of the yeast phagosomes (previously unlabeled) rapidly became colored. We inferred that this transfer was from endosomes (by Ph-E fusion) because Ph-L passage was blocked (by the NH4Cl). We conclude that NH4Cl induces Ph-E fusion at the same time as it suppressed Ph-L fusion. We discuss the mechanisms of these concurrent effects and suggest that they are independent; and we consider the implications of NH4Cl opening a direct route for endocytosed molecules to reach an intraphagosomal infection without involving lysosomes.

Ammonium Chloride↗
PubMed1991

Regulation of protein kinase A activation and prostaglandin E2-stimulated migration of Lewis lung carcinoma clones.

Lewis lung carcinoma (LLC) clones were used in in vitro models for dissemination to identify mechanisms regulating the stimulation of metastatic LLC-LN7 migration by prostaglandin E2 (PGE2) or forskolin plus 3-isobutyl-I-methylxanthine (IBMX), and the lack of responsiveness to generated cAMP in non-metastatic LLC-C8 cells. The regulatory subunits of protein kinase A (PKA) from LLC-LN7 cells bound more 8-N3-32P-cAMP, even though production of regulatory subunits was equal to that in LLC-C8 cells. Protein kinase C (PKC) differentially regulated PKA activation in the LLC variants. PKC activation inhibited PGE2-stimulated migration by LLC-LN7 cells. Inhibition of PKC with staurosporine stimulated LLC-LN7 cell migration to a level comparable with that induced by PGE2. However, PGE2 did not further stimulate the migration of staurosporine-treated cells. The PGE2 or staurosporine stimulation of LLC-LN7 cell migration was dependent on PKA activation. The effects that modulation of PKA and PKC had on LLC-LN7 cell migration paralleled the effects on endogenous protein phosphorylation. LLC-LN7 cell autophosphorylation was stimulated to a similar degree by PGE2, forskolin plus IMBX, staurosporine, or the combination of staurosporine and forskolin plus IBMX. In contrast, neither migration nor autophosphorylation was stimulated in non-metastatic LLC-C8 cells by cAMP elevation or by PKC inhibition. Autophosphorylation, although not migration, of LLC-C8 cells was stimulated by forskolin plus IBMX when PKC activity was inhibited. These results suggest that the increased PKA response of metastatic LLC-LN7 cells is contributed by an increased binding of cAMP by the PKA regulatory subunits and a reduced level of regulation by PKC.

Animals↗
PubMed1991

Tumoricidal activity of alveolar and peritoneal macrophages of C57BL/6 mice bearing metastatic or nonmetastatic variants of Lewis lung carcinoma.

The spontaneous tumoricidal abilities of alveolar and peritoneal macrophages from C57B1/6 mice bearing a metastatic or a nonmetastatic cloned variant of Lewis lung carcinoma (LLC) were measured. Cytotoxicity by alveolar macrophages was enhanced during the first few weeks after subcutaneous (s.c.) or intravenous (i.v.) injection of metastatic LLC-C3 cells, but not after injection of nonmetastatic LLC-C8 cells. Alveolar macrophages from mice with s.c.-injected metastatic tumors, but not with nonmetastatic tumors, could be further activated in vitro, but not beyond the maximal level of spontaneous cytotoxicity. Late in tumor growth, the spontaneous cytotoxicity by alveolar macrophages of metastatic LLC-C3 tumor bearers was suppressed and could not be increased by in vitro activation. The tumoricidal abilities of peritoneal macrophages from mice bearing either LLC-C3 or LLC-C8 tumors were modulated in a similar way, as were alveolar macrophages. The reduced cytotoxicity by alveolar macrophages from mice with nonmetastatic tumors or from mice bearing large metastatic tumors was not due to suppression by macrophage-derived prostaglandins. The loss of tumoricidal capabilities by macrophages from mice with large metastatic LLC-C3 tumors was not caused by elevated systemic prostaglandin E2 (PGE2) levels. These results suggest that alveolar and peritoneal macrophages are activated to be cytotoxic during development of pulmonary metastases and do not need to be functionally depressed for successful establishment of metastases.

Animals↗
PubMed1991

Antibodies to colony-stimulating factors block Lewis lung carcinoma cell stimulation of immune-suppressive bone marrow cells.

Progressive growth of metastatic Lewis lung carcinoma (LLC) tumors results in a concurrent stimulation of myelopoiesis and the appearance of immune-suppressive bone marrow cells. The present study has shown that normal bone marrow cells could be induced to become immune-suppressive by 3 days of culture with supernatants of cloned metastatic LLC-LN7 variant cells. The capacity of the LLC-LN7 supernatants to stimulate the appearance of suppressor cells was directly proportional to the concentration of supernatant used in the bone marrow culture. When adoptively transferred with a LLC-LN7 tumor inoculum, the supernatant-induced suppressor bone marrow cells increased the rate of appearance of palpable tumors and the frequency of tumor establishment. The LLC-LN7 supernatants containing suppressor-cell-inducing activity also had colony-stimulating factor (CSF) activity. The CSF activity produced by the LLC-LN7 cells could be diminished with neutralizing antibodies to either granulocyte/monocyte(GM-) CSF or to interleukin-3 (IL-3). Likewise, the suppressor-inducing activity in the LLC-LN7 supernatants was diminished by pretreatment with anti-GM-CSF or anti-IL-3. The combination of anti-GM-CSF and anti-IL-3 completely neutralized all suppressor-inducing activity produced by the LLC-LN7 cells. These results suggest that the secretion of IL-3 and GM-CSF by LLC-LN7 tumor cells is a mechanism by which the tumors stimulate myelopoiesis and induce normal bone marrow cells to become immune-suppressive. Bone marrow cells that are induced to become immune-suppressive by culture with LLC-LN7 supernatants can, in turn, facilitate the establishment of tumor in vivo.

Animals↗
PubMed1991

Effects of dietary fish oil on the induction of experimental membranous nephropathy in the rat.

We examined the effect of a fish oil-enriched diet on the development of experimental membranous nephropathy in the rat induced by administration of cationic bovine gamma globulin (CBGG). Rats were placed on a fish oil-enriched diet and control rats received a diet containing an equivalent amount of beef tallow. After 6 weeks on either diet, rats were pre-immunized and injected with CBGG. Proteinuria was significantly reduced in the fish oil-fed group as compared to the control group (160 +/- 40 mg/24 hours, n = 15, versus 280 +/- 36 mg/24 hours, n = 17, p less than 0.02). Glomerular filtration rate was also significantly higher in the fish oil-fed rats than in controls (0.91 +/- 0.07 ml/minute, n = 11, versus 0.60 +/- 0.05 ml/minute, n = 10, p less than 0.005). Glomerular production of prostaglandin E2 and thromboxane B2 the stable product of thromboxane A2, were inhibited by 68% and 70%, respectively, by the fish oil-enriched diet (n = 8, p less than 0.01 versus control). Glomerular leukotriene B4 was also inhibited by 50% in the fish oil-treated rats (n = 6, p less than 0.01), but inhibition of leukotriene B4 by the specific inhibitor L-663,536 in control rats did not ameliorate proteinuria. There was no difference in the amount of distribution of glomerular immune deposits as demonstrated by immunofluorescence and electron microscopy between the experimental and control groups. Moreover, comparable amounts of glomerular IgG deposits were present in the two groups. The specific immune response, assessed by measuring anti-BGG antibody levels, was not different between the two dietary groups, while more than 85% suppression of the splenic T- and B-cell mitogenic response to concanavalin-A and lipopolysaccharide was noted in rats fed the fish oil-enriched diet. We conclude that a fish oil-enriched diet reduces proteinuria and preserves the glomerular filtration rate in rats with CBGG-induced membranous nephropathy. Its mechanism of action remains to be established.

Animals↗
PubMed1991

Association of increased tumor cell responsiveness to prostaglandin E2 with more aggressive tumor behavior.

Secretion of prostaglandin E2 (PGE2) by cloned variants of QR fibrosarcoma or of Lewis lung carcinoma (LLC) did not correspond with their in vivo aggressive behavior. However, aggressiveness may be influenced by tumor responsiveness to PGE2. Each of the metastatic LLC or progressor QRpP variants was more motile in an in vitro migration model than were either the nonmetastatic LLC or regressor QR variants. One of the two tested progressor QR variants and all of the metastatic LLC variants were also more responsive to PGE2 in their in vitro migration through a membrane than were either regressor QR or nonmetastatic LLC variants. Thus, responsiveness to PGE2 by tumors may regulate the degree of tumor aggressiveness in vivo.

Animals↗